Bafilomycin A1 inhibits proteolytic cleavage and hydration but not yolk crystal disassembly or meiosis during maturation of sea bass oocytes

Oocytes of the black sea bass, Centropristes striata, were enlarged in volu me more than three-fold over a 24-hr period during oocyte maturation, both in vivo and in vitro. At the same time, the opaque oocytes clarified while the crystalline yolk inclusions lost their ordered structure, fused with on e another, and formed a continuous electron-lucent mass. The oocyte size in crease was due almost entirely to water uptake, which was accompanied by th e accumulation of Na+, K+, and free amino acids (FAAs). The absolute amount s of each of these small molecular weight osmotic effectors increased 2x, 4 x, and over 10x, respectively, indicating that the generation of FAAs is th e major cause of water uptake during maturation. Amino acid analyses indica ted that the amounts of all amino acids except taurine increased, so that s elective amino acids were not produced during maturation. The increase in F AAs was accompanied by the loss of certain high-molecular-weight yolk prote ins and the generation of many smaller peptides. Oocytes stimulated to unde rgo maturation in the presence of bafilomycin A1, a specific inhibitor of t he vacuolar ATPase-dependent proton pump, clarified and underwent maturatio n but did not increase significantly in size. Cytological examination revea led that yolk crystals fused and became homogeneous but maintained their el ectron density. No evidence of proteolysis was found in bafilomycin A1-trea ted oocytes and the generation of FAAs together with hydration was inhibite d in a dose-dependent manner (I-50 = 3 nM bafilomycin A1). Taken together, we postulate that the pronounced oocyte hydration in marine teleosts that s pawn pelagic (floating) eggs is accomplished by a two-step process whereby (i) K+ influx promotes yolk crystal disassembly and yolk sphere fusion and (ii) acidification of the yolk spheres activates yolk proteolysis and conco mitant hydration. Bafilomycin A1 inhibits only the second step so that many of the events of oocyte maturation, including germinal vesicle breakdown, occur in its presence but oocyte hydration is suppressed. (C) 2001 Wiley-Li ss, Inc.